Diagnosis of allergies



fiatenterl Oct. 9, 1 951 DIAGNOSIS OF ALLERGIES William Holmes Gardner, West Palm Beach, Fla.'

N Drawing. Application April 8, 1948,

Serial No. 19,892

2 Claims. (01.161 7 The present invention relates to the diagnosis of allergies in humanbeings, and has particular reference to a diagnostic compound and a new method for forming a preparation for diagnosing allergies in patients by determining patient reaction to allergenic substances under question in terms of action of the allergenic substances on the blood of the patient.

, An object of the invention is to provide a novel diagnostic preparation for the purpose of making rapid andefficient determination of the reaction of a patient to various allergenic substances. V

- A further object is to provide a novel method for the determination of allergies by the administration of specific allergenic substances to the patient intradermally.

A further object is to provide a novel method for the determination of allergies in human beings by the use of blood samples, whereby the discomfort to the patient attendant the commonly practiced skin transfer test methods is eliminated.

A further object is to provide a method of testing for allergies whereby a series of tests for determining patient reactions to a plurality of different allergenic substances may be accomplished simultaneously, without resort to the commonly known scratch test where the skin of the patient is broken for the introductio of suspect allergenic substances. l

A further object is to provide a novel method of testing for allergenic reaction wherein the results of the tests may be speedily and accurately determined in a manner permitting of accurate analysis and determination of a relative degree of patient sensitivity to the allergenic substance in question. I

' A further object is toprovide a novel metho for forming preparations for use in the diagnosis of allergies. V A

'Yet a furtherobject is to provide an agglutination test for the determination of allergies in human beings whereby the presence of sensitivity to certain substances such as foods, pollens, plants, inhalants and the like, may be determined by laboratory methods.

In accordance with the invention, the diagnostic preparations employed comprise cultures of colon bacillus which are specific to the foods, pollens, plants, inhalants, and the like which may comprise the source of the disturbance in the patient. The cultures may be employed to determine the presence or absence of any sensitivity in the patient to the specific colon bacillus.

employed in forming the cultures.

In preparing the cultures, a specific colon bacillus isolated fromsuspeeted sources of allergenic disturbances, such as a particular animal, plant, grass, tree, food, etc. 7

The Bacillus coli employed in the constitution of a specific diagnostic preparation may be found in any portion of a plant. tree, animal, or other substance in question. For example, when the suspect substance comprises pork or other flesh of the pig or hog, Bacillus coli specific to the source may be foundin the flesh of the hog.

In some instances more readily available or prolific sources of Bacillus 00 may be found by investigation of the suspect substance. In the case of animal meats, for example, the liver was found to be the most prolific source of Bac'illus coli. Inthe caseof plants, the Bacillus coli may be found in the fruit of the plant, the most prolific source comprising fruit in the green or unmatured. state, and new growths on the plant. The'same is true for weeds and grasses.

In the case of trees, such as for examplethe orange or apple tree, Bacillus coli specific to these sources may be found in the green fruit, the most prolific source being newly formed green shoots containing a high concentration of the sap of the tree.

The pure culture is then incubated in a media suitable for growing colon bacilli. As an example of a specific preparation, where pork is the suspect substance, a small section of fresh hog liver is dropped into a standard media for growing Bacillus coli such as a'sterile beef-peptonelactose broth. The incubation in this broth may proceed for a sufiicienttime to procure an active, concentrated culture; as for example, a period of forty-eighthours at 38 C. Incubation may be accelerated or extended as desired in accordance with known laboratory techniques including variations in temperature, humidity, and the incubation media employed.

The culture is then inactivated, standardized and diluted, according to usual laboratory methods governing the preparation of culture for bacterial agglutination tests, such as the commonly known Widal test for typhoid. For example, a smear culture of the broth prepared above is grown on sterile agar plates, being incubated for a suitable period, for example twenty-four to thirty-six hours at 38 C. Colonies of Bacillus coli developed are isolated and identified as such by comparison with known cultural characteristics, specific media, and microscopic examina-' equal amounts of saline solution, and is then ready for use. Greater dilutionsmaybedesiv able in certain instances.

The culture thus prepared, contains a colon bacillus having properties specific to the cultures prepared from the particular source, and thus permits of specific diagnosis of allergies based upon antibody -reactions occur'ri-ng in "the blood of the patient. r

-A-markedadvantage of -the 'diagnosticrpreparations-of the invention resides --'in their adaptivity to use in blood agglutination tests, whereby all skin testing is eliminated :and a llergies may be determined 'by laboratory methods alone. In this case, the specific colon *bac-illus iculture is .=combined with the blood =serum-lof the patient whose allergies are to be determined. For examplepone drop of the blood serum-.of the-patient may he added to =nedr0p of theBacillus colz' sohition, placed in a sterile test tub and incubated for twelve hours. The-solution is then dropped onto a (glass slide and read microscopically .to :determine-if agglutination has occurred, indicating a positivereactionto the Bacillus .coli :of the suspect substance. The degree :of hypersensitivity of the patient :to thesubstance is determined .by the .degree ofragglutination. or-lantibody; reaction.

.It .is preferred to standardize theedegrees vof agglutination termsof arbitrary meanings ref one, two, three .orl'four eplus similar :to the treadingsof .the well known Widal test .=or typh i permitting. of a standardizedncomplete diagnosis of a particular allergy based upon anti-body reaction. H V

In carrying out the agglutination tests, .blood serum of the patient must be employed. Whole blood or hemolized blood cannot be .used.

The cultures prepared in accordance with the invention mayalsobe adminis'teredito the patient intradermally. 'In .such case, the cultures prepared and treated as above described preferably are filtered prior to injection between thelayers of skin ofthe patient. Sensitivity of'the patient to the specific colonbac'illus or a positive reaction'to the test, is shown by the usual" skin reaction to 'intradermal tests, -negative reactions showing no evidence of irritation or'other skin disturbance. I I

Use of specific cultures of colon bacillus "in the determination of allergies, both by intra dermal application and in bacterial agglutination tests, has been thoroughly checked for accuracy of result against the accepted passive transfer tests and clinically, as well as against each other. The results in all tests were found to coincide completely in terms of positive diagnosis and subsequent tests of suspect substances found to be positive against *clinical symptoms iappearing upon patient exposure.

The principles of the invention as set forth are capable of adaptation in accordance with known procedures :by those familiar with this field, and accordingly the invention is to be limited only as 'defined'in'the appended claims.

.Iclaim:

..1. A method of determining sensitivity of the patient to a suspect source of allergenic disturbance comprising isolating B. coll taken from the suspect source, culturing the B. coll from the suspect source to =provide a colonyof B. =c'0li,.iinadtivating the culture, and diluting the inactivated culture withsterilesalinesolution to make anextract having the allergenic characteristic -of the suspect source, mixing the extract-with-tl=re bloo'd serum o'f the patient-toincur agglutination in-de gree proportional -to the --degr-ee-o f sensitivity of the patient to the suspect -source,-an'd observing the agglutination #of the mi x'ture to determine the sensitivity of the -pa=tiento the source of the B.-coli.

"-2. A method of -aeterm'ini-ngallergenic sensitivity of a patient to a source of allergenic-disturbance comprising iso'la-ting co'li taken from the suspect-source, "culturing the 12.com "from the suspect source to -previde a -colony of B. co'li, inactivating the culture, "anddiluting the =inactivated culture with-sterile -saline solution ta'make an extract having the allergenic characteristic of the suspect source, withdrawing a samplewf blood from the =patient,-separating the blood serum from the remainder-or the blood in the sample, and mixing the extract with tlie bleod serum, and :observing the --'aggluti-nationof the mixture to determine ithe-sensitivity of the patient to thesource of theiB.:coli.

Zinsser: Textbook of Bacteriology1".7th ed, 1935,pp.5'53 560.

Newand"NonoflicialRemedies, 1945 (Am. "Med. Assn.) ,Pp. 42-46.

Number 

1. A METHOD OF DETERMINING SENSITIVITY OF THE PATIENT TO A SUSPECT SOURCE OF ALLERGENIC DISTURBANCE COMPRISING ISOLATING B. COLI TAKEN FROM THE SUSPECT SOURCE, CULTURING THE B. COLI FROM THE SUSPECT SOURCE TO PROVIDE A COLONY OF B. COLI, INACTIVATING THE CULTURE, AND DILUTING THE INACTIVATED CULTURE WITH STERILE SALINE SOLUTION TO MAKE AN EXTRACT HAVING THE ALLERGIC CHARACTERISTIC OF THE SUSPECT SOURCE, MIXING THE EXTRACT WITH THE BLOOD SERUM OF THE PATIENT TO INCUR AGGLUTINATION IN DEGREE PROPORTIONAL TO THE DEGREE OF SENSITIVITY OF THE PATIENT TO THE SUSPECT SOURCE, AND OBSERVING THE AGGLUTINATION OF THE MIXTURE TO DETERMINE THE SENSITIVITY OF THE PATIENT TO THE SOURCE OF THE B. COLI. 